Choosing the Method of Crystallization to Obtain Optimal Results

Govada, Lata; Chayen, Naomi E.

doi:10.3390/cryst9020106

Cited by 8 publications

(9 citation statements)

References 34 publications

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“…Nevertheless, vapor diffusion still remains an efficient method to screen 2 of 13 crystallization conditions using a low quantity of protein. Regardless of the method, crystallization is potentially able to reduce downstream manufacturing costs by improving the efficiency of the protein purification step with respect to conventional chromatographic methods [5][6][7][8][9][10][11]. Moreover, the active ingredient is usually more stable in crystal form than in solution, a feature that is crucial in the case of samples that easily degrade, such as protein molecules.…”

Section: Introductionmentioning

confidence: 99%

Optimization of Vapor Diffusion Conditions for Anti-CD20 Crystallization and Scale-Up to Meso Batch

Yang

Belviso

et al. 2019

Crystals

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The crystal form is one of the preferred formulations for biotherapeutics, especially thanks to its ability to ensure high stability of the active ingredient. In addition, crystallization allows the recovery of a very pure drug, thus facilitating the manufacturing process. However, in many cases, crystallization is not trivial, and other formulations, such as the concentrate solution, represent the only choice. This is the case of anti-cluster of differentiation 20 (anti-CD20), which is one of the most sold antibodies for therapeutic uses. Here, we propose a set of optimized crystallization conditions for producing anti-CD20 needle-shaped crystals within 24 h in a very reproducible manner with high yield. High crystallization yield was obtained with high reproducibility using both hanging drop vapor diffusion and meso batch, which is a major step forward toward further scaling up the crystallization of anti-CD20. The influence of anti-CD20 storage conditions and the effect of different ions on the crystallization processes were also assessed. The crystal quality and the high yield allowed the first crystallographic investigation on anti-CD20, which positively confirmed the presence of the antibody in the crystals.

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Section: Introductionmentioning

confidence: 99%

Optimization of Vapor Diffusion Conditions for Anti-CD20 Crystallization and Scale-Up to Meso Batch

Yang

Belviso

et al. 2019

Crystals

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“…Fourteen protomers of XoGroEL constitute a large complex of more than 800 kDa. Although various approaches for the crystallization of a mega Dalton protein complex have been developed [23,24], the determination of the crystal structure of such a large complex at high quality is still challenging. The overall B factor (temperature factor) of XoGroEL is 82.3.…”

Section: Discussionmentioning

confidence: 99%

Crystal Structure of Chaperonin GroEL from Xanthomonas oryzae pv. oryzae

et al. 2019

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Xanthomonas oryzae pv. oryzae (Xoo) is a plant pathogen that causes bacterial blight of rice, with outbreaks occurring in most rice-growing countries. Thus far, there is no effective pesticide against bacterial blight. Chaperones in bacterial pathogens are important for the stabilization and delivery of effectors into host cells to cause disease. In bacteria, GroEL/GroES complex mediates protein folding and protects proteins against misfolding and aggregation caused by environmental stress. We determined the crystal structure of GroEL from Xanthomonas oryzae pv. oryzae (XoGroEL) at 3.2 Å resolution, which showed the open form of two conserved homoheptameric rings stacked back-to-back. In the open form structure, the apical domain of XoGroEL had a higher B factor than the intermediate and equatorial domains, indicating that the apical domain had a flexible conformation before the binding of substrate unfolded protein and ATP. The XoGroEL structure will be helpful in understanding the function and catalytic mechanism of bacterial chaperonin GroELs.

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“…Once a critical nucleus is formed, growth follows spontaneously [56][57][58][59]. Nonetheless, excess nucleation consistently occurs, resulting in the formation of numerous low-quality micro-/nano-crystals [60,61].…”

Section: Polycrystalline Samples and First Virus Protein Xrpd Studiesmentioning

confidence: 99%

Applications of X-ray Powder Diffraction in Protein Crystallography and Drug Screening

et al. 2020

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Providing fundamental information on intra/intermolecular interactions and physicochemical properties, the three-dimensional structural characterization of biological macromolecules is of extreme importance towards understanding their mechanism of action. Among other methods, X-ray powder diffraction (XRPD) has proved its applicability and efficiency in numerous studies of different materials. Owing to recent methodological advances, this method is now considered a respectable tool for identifying macromolecular phase transitions, quantitative analysis, and determining structural modifications of samples ranging from small organics to full-length proteins. An overview of the XRPD applications and recent improvements related to the study of challenging macromolecules and peptides toward structure-based drug design is discussed. This review congregates recent studies in the field of drug formulation and delivery processes, as well as in polymorph identification and the effect of ligands and environmental conditions upon crystal characteristics. These studies further manifest the efficiency of protein XRPD for quick and accurate preliminary structural characterization.

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Choosing the Method of Crystallization to Obtain Optimal Results

Cited by 8 publications

References 34 publications

Optimization of Vapor Diffusion Conditions for Anti-CD20 Crystallization and Scale-Up to Meso Batch

Optimization of Vapor Diffusion Conditions for Anti-CD20 Crystallization and Scale-Up to Meso Batch

Crystal Structure of Chaperonin GroEL from Xanthomonas oryzae pv. oryzae

Applications of X-ray Powder Diffraction in Protein Crystallography and Drug Screening

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